Method for preparing flavor peptide through colloid mill-high-pressure microjet homogenization combined auxiliary enzymolysis and application of flavor peptide

Abstract

The invention discloses a method for preparing flavor peptide through colloid mill-high-pressure microjet homogenization combined auxiliary enzymolysis and application, and belongs to the technical field of flavor peptide preparation. According to the method, cells of the edible mushrooms are broken and dissolved through a colloid mill-high-pressure microjet homogenization combined technology, so that the particle size of the materials can reach the nanometer level, protein in the materials can be degraded by enzymes more easily, the enzymolysis efficiency can be remarkably improved, and a traditional first step of cellulase enzymolysis wall breaking is omitted; the prepared flavor peptide has better quality, flavor and taste; compared with the prior art, no chemical reagent or solvent needs to be added, pollution to the environment is reduced, meanwhile, energy consumption needed by the technology is relatively low and is 2433.6 + / -487 kJ / h, the release rate of soluble nitrogen reaches up to 81.66 + / -0.96%, the material loss rate is 2.37 + / -0.14%, the production cost can be reduced, and the requirement for sustainable development is met.

Description

Technical Field

[0001] The invention relates to the technical field of flavor peptide preparation, in particular to a method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis and its application. Background Art

[0002] For the preparation of edible fungus flavor peptides, the traditional water extraction method does not pay enough attention to the cell wall breaking process. Enzymatic cell wall breaking is widely used due to its mild conditions and strong specificity. However, the enzymes currently used are of single variety and take a long time, which often leads to the growth of microorganisms. Therefore, the use of physical cell wall breaking has become a reasonable trend.

[0003] High-pressure microfluidization technology is a good way to physically break the wall. It is a new type of homogenization method that can be operated continuously, and can achieve a series of operations such as mixing, ultrafine grinding, pressurization, and homogenization of the material in a short time. By utilizing the high pressure generated by the hydraulic pump, the fluid is dispersed into two or more strands in the microchannel, and the thin stream undergoes a strong high-pressure collision in the impact chamber. During the collision, a huge pressure drop is generated, causing the liquid to undergo a comprehensive effect such as high-speed collision, high-frequency vibration, high-frequency shear, cavitation, and instantaneous pressure drop. In the process of sample processing, the material is simultaneously subjected to multiple mechanical forces and their thermal effects, thereby inducing changes in the structure and properties of biological macromolecules. Edible fungi are rich in cellulose, and the combination of colloid milling and high-pressure microfluidization homogenization can achieve the best wall-breaking effect in the enzymatic hydrolysis of edible fungi.

[0004] At present, my country is the world's largest producer, consumer and exporter of edible fungi, with rich edible fungi resources. With the development of social economy, people tend to eat healthier and pursue more and more natural umami formulas. Flavor peptides are a kind of fresh-tasting substances, mainly derived from animals and plants. Compared with traditional umami seasonings, they have a soft taste, certain functional characteristics and processing adaptability. Flavor peptides can increase the flavor and umami of food, and can play a synergistic role in enhancing the freshness of other substances, thereby reducing the body's intake of salt and monosodium glutamate. Edible fungi are rich in nutrients, have a unique fresh fragrance and contain more fresh substances. Edible fungi flavor peptides can be extracted and prepared by enzymatic hydrolysis, thus providing a theoretical basis for the development of natural seasonings. Summary of the invention

[0005] The purpose of the present invention is to provide a method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis and its application, so as to provide a method for preparing flavor peptides with low energy consumption, low cost, high preparation efficiency, low environmental pollution and better quality of prepared flavor peptides.

[0006] To achieve the above object, the present invention provides a method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis, the steps of which are as follows:

[0007] 1) After the dried edible fungi are crushed, water 10-30 times the mass of the dried powder is added, and the mixture is beaten with a colloid mill for 5-10 minutes, and then homogenized with a high-pressure microfluidizer to obtain an initial homogenate of the edible fungi for later use;

[0008] 2) adjusting the pH value, adding flavor protease, performing enzymatic hydrolysis, and then performing enzyme inactivation treatment in boiling water;

[0009] 3) adjusting the pH value again, adding xylose and lysine, and performing the Maillard reaction;

[0010] 4) cooling the obtained edible fungus Maillard liquid to room temperature, and filtering out the edible fungus residue to obtain a supernatant;

[0011] 5) pre-filtering the edible fungus Maillard liquid with an organic filter membrane, and then passing the edible fungus Maillard liquid through an ultrafiltration membrane with a molecular weight cutoff of >10 kDa, 5k-10 kDa, 3k-5 kDa and <3 kDa to obtain four components with different molecular weights;

[0012] 6) spray drying the filtrate to obtain edible fungus flavor peptides of four components, namely, >10 kDa, 5k-10 kDa, 3k-5 kDa and <3 kDa, and performing sensory evaluation on the flavor peptides of different components;

[0013] 7) Select the target peptide, and package the obtained target peptide using a heat shrink film packaging machine at room temperature.

[0014] Preferably, in step 1), the pitch gap of the colloid mill is 100-200 μm, and the pressure parameter of the high-pressure microfluidizer is 160-200 MPa, so that the particle size of the edible fungi is no more than 100 nm.

[0015] Preferably, in step 2), the pH value is adjusted to 6-7, the amount of flavor protease added is 2-4wt% of the dry matter of the edible fungi, the enzymolysis time is 2-3h, and after the enzymolysis is completed, the enzyme is inactivated in boiling water for 15min.

[0016] Preferably, in step 3), Maillard reaction is carried out, the reaction pH is 6.5-8, the reaction temperature is 90-120° C., the reaction time is 90-120 min, the amount of xylose added is 10-12 wt % of the dry matter of the edible fungus, and the amount of lysine added is 6-8 wt % of the dry matter of the edible fungus.

[0017] Preferably, four layers of gauze are used for filtering in step 4).

[0018] Preferably, in step 5), a 0.45 μm organic filter membrane is used to pre-filter the sample solution.

[0019] Preferably, in step 6), the spray drying parameters of the filtrate are as follows: air volume of 70%, temperature of 180° C., and feed rate of 30%, and the flavor peptides of different components are evaluated for umami taste using a sensory evaluation standard table.

[0020] Sensory evaluation standard table refers to Xiao Chen, Jingyang Yu, Heping Cui, et al. Effect of Temperature on Flavour Compounds and Sensory Characteristics of Maillard Reaction on Products Derived form Mushroom Hydrolysate[J]. Molecules, 2018, 23: 247.

[0021] The invention discloses an application of the method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis in the preparation of flavor peptides.

[0022] Therefore, the method and application of preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis provided by the present invention have the following specific technical effects:

[0023] (1) The present invention uses a colloid mill-high-pressure microfluidization homogenization combined technology to break and dissolve edible fungi cells, which can make the particle size of the material reach the nanometer level, making the protein inside the material easier to be degraded by enzymes, thereby significantly improving the enzymatic hydrolysis efficiency and omitting the traditional first step of cellulase enzymatic hydrolysis and cell wall breaking;

[0024] (2) The flavor peptide product prepared by the process provided by the present invention has better quality, flavor and taste. The traditional method for preparing flavor peptides of edible fungi usually requires a long enzymatic hydrolysis time, while the colloid mill-high pressure microfluidization homogenization combined technology can significantly shorten the enzymatic hydrolysis time and accelerate the wall breaking speed of edible fungi, which can not only improve production efficiency, but also save time and cost;

[0025] (3) The process provided by the present invention does not require the addition of any chemical reagents or solvents, thereby reducing environmental pollution. At the same time, the energy consumption required by the technology is relatively low, which is 2433.6±487 kJ / h. The soluble nitrogen release rate is as high as 81.66±0.96%, and the material loss rate is 2.37±0.14%, which helps to reduce production costs and meets the requirements of sustainable development.

[0026] The technical solution of the present invention is further described in detail below through the accompanying drawings and embodiments. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings required for use in the description of the embodiments of the present invention will be briefly introduced below. Obviously, the accompanying drawings in the following description are only some embodiments of the present invention. For ordinary technicians in this field, other accompanying drawings can be obtained based on these accompanying drawings without paying creative labor.

[0028] Figure 1 It is a sensory evaluation radar chart of three kinds of mushrooms (A), Agrocybe edulis (B), Cordyceps sinensis (C), Boletus edulis (D), Pleurotus eryngii (E), and Pleurotus eryngii (F) tested in the present invention;

[0029] Figure 2 It is a flow chart of the preparation method provided by the present invention. DETAILED DESCRIPTION

[0030] The technical solution of the present invention is further described below through the accompanying drawings and embodiments.

[0031] In order to make the purpose, technical scheme and advantages of the present application clearer, more thorough and more complete, the technical scheme of the present invention is clearly and completely described below through the accompanying drawings and examples. The following detailed descriptions are all descriptions of the embodiments, and are intended to provide further detailed descriptions of the present invention. Unless otherwise specified, all technical terms used in the present invention have the same meaning as those generally understood by those skilled in the art to which the present application belongs.

[0032] The instruments, equipment, reagents and materials used in the examples were all obtained through commercial channels.

[0033] Embodiment 1

[0034] The shiitake mushroom flavor peptides were prepared by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis. Figure 2 As shown, the specific steps are as follows:

[0035] (1) After the dried shiitake mushrooms were crushed by a high-speed crusher, 40 g was weighed, 800 mL of pure water (solid-liquid ratio was 1:20) was added, and the mixture was slurried by a colloid mill for 8 min. The high-pressure microfluidizer was set to 180 MPa for homogenization to obtain a homogenate.

[0036] (2) The pH value of the slurry was adjusted to 6.5 with 1M HCl and 1M NaOH, and 0.076 g (4 wt%) flavor protease (20000 U / g) was added, and the slurry was enzymatically hydrolyzed at 50°C for 2 h. After the enzymatic hydrolysis, the slurry was placed in boiling water for 15 min to inactivate the enzyme.

[0037] (3) The pH value of the solution was then adjusted to 7 with 1 M HCl and 1 M NaOH, 0.19 g (10 wt %) of xylose and 0.114 g (6 wt %) of lysine were added, and Maillard reaction was carried out at 100° C. for 100 min.

[0038] (4) Cooling the shiitake mushroom Maillard liquid obtained in step (3) to room temperature, filtering out the edible mushroom residue with four layers of gauze to obtain a supernatant.

[0039] (5) The obtained supernatant was pre-filtered using a 0.45 μm organic filter membrane, and the sample solution obtained by pre-filtration was then passed through an ultrafiltration membrane with a molecular weight cutoff of >10 kDa, 5k-10 kDa, 3k-5 kDa and <3 kDa to obtain four components with different molecular weights.

[0040] (6) The final filtrate is spray-dried to obtain four components of shiitake mushroom flavor peptides, namely, >10 kDa, 5k-10 kDa, 3k-5 kDa, and <3 kDa, and sensory evaluation is performed on the shiitake mushroom flavor peptides of different components.

[0041] (7) Selecting the target peptide, and packaging the obtained target peptide using a heat shrink film packaging machine at room temperature.

[0042] Embodiment 2

[0043] The tea tree mushroom flavor peptide was prepared by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis. The specific steps are as follows:

[0044] (1) After the dried Agrocybe aegerita was crushed by a high-speed crusher, 40 g was weighed, 1200 mL of purified water (solid-liquid ratio was 1:30) was added, and the mixture was slurried by a colloid mill for 10 min. The high-pressure microfluidizer was set to a homogenization pressure of 200 MPa to obtain a homogenate.

[0045] (2) The pH value of the slurry was adjusted to 7 with 1M HCl and 1M NaOH, and 0.052 g (4 wt%) flavor protease (20000 U / g) was added, and the slurry was enzymatically hydrolyzed at 55°C for 3 h. After the enzymatic hydrolysis, the slurry was placed in boiling water for 15 min to inactivate the enzyme.

[0046] (3) The pH value of the solution was then adjusted to 7.5 with 1 M HCl and 1 M NaOH, and 0.156 g (12 wt %) of xylose and 0.078 g (6 wt %) of lysine were added, followed by Maillard reaction at 110° C. for 100 min.

[0047] (4) Cooling the Agrocybe aegerita Maillard liquid obtained in step (3) to room temperature, filtering out the edible fungus residue using four layers of gauze to obtain a supernatant.

[0048] (5) The obtained supernatant was pre-filtered using a 0.45 μm organic filter membrane, and the sample solution obtained by pre-filtration was then passed through an ultrafiltration membrane with a molecular weight cutoff of >10 kDa, 5k-10 kDa, 3k-5 kDa and <3 kDa to obtain four components with different molecular weights.

[0049] (6) The final filtrate is spray-dried to obtain four components of Agrocybe aegerita flavor peptides, namely, >10 kDa, 5k-10 kDa, 3k-5 kDa, and <3 kDa. Sensory evaluation is performed on the Agrocybe aegerita flavor peptides of different components.

[0050] (7) Selecting the target peptide, and packaging the obtained target peptide using a heat shrink film packaging machine at room temperature.

[0051] Embodiment 3

[0052] The cordyceps flower flavor peptide was prepared by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis, and the specific steps are as follows:

[0053] (1) After the dried Cordyceps flower is crushed with a high-speed crusher, 40 g is weighed, 600 mL of pure water (solid-liquid ratio is 1:15) is added, and the mixture is slurried with a colloid mill for 5 min. The high-pressure microfluidizer is set to a homogenization pressure of 160 MPa to obtain a homogenate.

[0054] (2) The pH value of the slurry was adjusted to 6.5 with 1M HCl and 1M NaOH, and 0.1 g (4 wt%) flavor protease (20000 U / g) was added, and the slurry was enzymatically hydrolyzed at 50°C for 2 h. After the enzymatic hydrolysis, the slurry was placed in boiling water for 15 min to inactivate the enzyme.

[0055] (3) The pH value of the solution was then adjusted to 7 with 1 M HCl and 1 M NaOH, 0.25 g (10 wt %) xylose and 0.15 g (6 wt %) lysine were added, and Maillard reaction was carried out at 90° C. for 100 min.

[0056] (4) Cooling the Cordyceps flower Maillard liquid obtained in step (3) to room temperature, filtering out the edible fungus residue using four layers of gauze to obtain a supernatant.

[0057] (5) The obtained supernatant was pre-filtered using a 0.45 μm organic filter membrane, and the sample solution obtained by pre-filtration was then passed through an ultrafiltration membrane with a molecular weight cutoff of >10 kDa, 5k-10 kDa, 3k-5 kDa and <3 kDa to obtain four components with different molecular weights.

[0058] (6) The final filtrate is spray-dried to obtain four components of Cordyceps flower flavor peptides: >10 kDa, 5k-10 kDa, 3k-5 kDa, and <3 kDa. The Cordyceps flower flavor peptides of different components are subjected to sensory evaluation.

[0059] (7) Selecting the target peptide, and packaging the obtained target peptide using a heat shrink film packaging machine at room temperature.

[0060] Embodiment 4

[0061] The boletus flavor peptide was prepared by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis, and the specific steps are as follows:

[0062] (1) After the dried boletus was crushed by a high-speed crusher, 40 g was weighed, 1000 mL of pure water (solid-liquid ratio was 1:25) was added, and the mixture was slurried by a colloid mill for 10 min. The high-pressure microfluidizer was set to a homogenization pressure of 180 MPa to obtain a homogenate.

[0063] (2) The pH value of the slurry was adjusted to 6.5 with 1M HCl and 1M NaOH, and 0.06 g (4 wt%) flavor protease (20000 U / g) was added, and the slurry was enzymatically hydrolyzed at 55°C for 3 h. After the enzymatic hydrolysis, the slurry was placed in boiling water for 15 min to inactivate the enzyme.

[0064] (3) The pH value of the solution was then adjusted to 7 with 1 M HCl and 1 M NaOH, and 0.18 g (12 wt %) of xylose and 0.12 g (8 wt %) of lysine were added, followed by Maillard reaction at 100° C. for 90 min.

[0065] (4) Cooling the porcini Maillard liquid obtained in step (3) to room temperature, filtering out the edible mushroom residue using four layers of gauze to obtain a supernatant.

[0066] (5) The obtained supernatant was pre-filtered using a 0.45 μm organic filter membrane, and the sample solution obtained by pre-filtration was then passed through an ultrafiltration membrane with a molecular weight cutoff of >10 kDa, 5k-10 kDa, 3k-5 kDa and <3 kDa to obtain four components with different molecular weights.

[0067] (6) spray drying the final filtrate to obtain boletus flavor peptides of four components: >10 kDa, 5k-10 kDa, 3k-5 kDa, and <3 kDa.

[0068] Sensory evaluation of different components of boletus flavor peptides was carried out.

[0069] (7) Selecting the target peptide, and packaging the obtained target peptide using a heat shrink film packaging machine at room temperature.

[0070] Embodiment 5

[0071] The colloid mill-high pressure microfluidizer homogenization combined with assisted enzymatic hydrolysis was used to prepare Pleurotus eryngii flavor peptides. The specific steps are as follows:

[0072] (1) After the dried Pleurotus eryngii was crushed by a high-speed crusher, 40 g was weighed, 800 mL of purified water (solid-liquid ratio was 1:20) was added, and the mixture was slurried by a colloid mill for 8 min. The high-pressure microfluidizer was set to a homogenization pressure of 180 MPa to obtain a homogenate.

[0073] (2) The pH value of the slurry was adjusted to 6.5 with 1M HCl and 1M NaOH, and 0.076g (4wt%) flavor protease (20000U / g) was added, and the slurry was enzymatically hydrolyzed at 50°C for 2h30min. After the enzymatic hydrolysis, the slurry was placed in boiling water for 15min to inactivate the enzyme.

[0074] (3) The pH value of the solution was then adjusted to 7 with 1 M HCl and 1 M NaOH, and 0.19 g (10 wt %) of xylose and 0.133 g (7 wt %) of lysine were added, followed by Maillard reaction at 110° C. for 90 min.

[0075] (4) Cooling the Pleurotus eryngii Maillard liquid obtained in step (3) to room temperature, filtering out the edible fungus residue with four layers of gauze to obtain a supernatant.

[0076] (5) The obtained supernatant was pre-filtered using a 0.45 μm organic filter membrane, and the sample solution obtained by pre-filtration was then passed through an ultrafiltration membrane with a molecular weight cutoff of >10 kDa, 5k-10 kDa, 3k-5 kDa and <3 kDa to obtain four components with different molecular weights.

[0077] (6) The final filtrate is spray-dried to obtain Pleurotus eryngii flavor peptides of four components, namely, >10 kDa, 5k-10 kDa, 3k-5 kDa, and <3 kDa. Sensory evaluation is performed on the Pleurotus eryngii flavor peptides of different components.

[0078] (7) Selecting the target peptide, and packaging the obtained target peptide using a heat shrink film packaging machine at room temperature.

[0079] Embodiment 6

[0080] The colloid mill-high pressure microfluidizer homogenization combined with assisted enzymatic hydrolysis was used to prepare the white jade mushroom flavor peptide. The specific steps are as follows:

[0081] (1) After the dried white jade mushrooms were crushed by a high-speed crusher, 40 g was weighed, 800 mL of purified water (solid-liquid ratio was 1:20) was added, and the mixture was slurried by a colloid mill for 10 min. The high-pressure microfluidizer was set to a homogenization pressure of 180 MPa to obtain a homogenate.

[0082] (2) The pH value of the slurry was adjusted to 7 with 1M HCl and 1M NaOH, and 0.076 g (4 wt%) flavor protease (20000 U / g) was added, and the slurry was enzymatically hydrolyzed at 50°C for 3 h. After the enzymatic hydrolysis, the slurry was placed in boiling water for 15 min to inactivate the enzyme.

[0083] (3) The pH value of the solution was then adjusted to 7.5 with 1 M HCl and 1 M NaOH, and 0.228 g (12 wt %) of xylose and 0.114 g (6 wt %) of lysine were added, followed by Maillard reaction at 100° C. for 115 min.

[0084] (4) Cooling the Pleurotus eryngii Maillard liquid obtained in step (3) to room temperature, filtering out the edible fungus residue with four layers of gauze to obtain a supernatant.

[0085] (5) The obtained supernatant was pre-filtered using a 0.45 μm organic filter membrane, and the sample solution obtained by pre-filtration was then passed through an ultrafiltration membrane with a molecular weight cutoff of >10 kDa, 5k-10 kDa, 3k-5 kDa and <3 kDa to obtain four components with different molecular weights.

[0086] (6) The final filtrate is spray-dried to obtain four components of Pleurotus eryngii flavor peptides, namely, >10 kDa, 5k-10 kDa, 3k-5 kDa, and <3 kDa. Sensory evaluation is performed on the Pleurotus eryngii flavor peptides of different components.

[0087] (7) Selecting the target peptide, and packaging the obtained target peptide using a heat shrink film packaging machine at room temperature.

[0088] Comparative Example 1

[0089] The specific steps for preparing flavor peptides of shiitake mushroom, Agrocybe oleracea, Cordyceps sinensis, Boletus edulis, Pleurotus eryngii and Pleurotus eryngii are exactly the same as those of Examples 1 to 6, except that step (1) is respectively:

[0090] Shiitake mushroom flavor peptide: After the dried shiitake mushrooms were crushed with a high-speed grinder, 40 g was weighed, 800 mL of pure water (solid-liquid ratio was 1:20) was added, and the mixture was pulped with a colloid mill for 8 min, and the high-pressure microfluidization homogenization pressure was set to 180 MPa for homogenization to obtain a homogenate. The pH value of the homogenate was adjusted to 4 with 1 M HCl and 1 M NaOH, 0.076 g (4 wt%) of cellulase (20,000 U / g) was added, and enzymolysis was carried out at 50°C for 2 h.

[0091] Agrocybe agrocybe flavor peptide: After the dried Agrocybe agrocybe was crushed with a high-speed pulverizer, 40 g was weighed, 1200 mL of pure water (solid-liquid ratio was 1:30) was added, and the mixture was slurried with a colloid mill for 10 min, and the high-pressure microfluidization homogenization pressure was set to 200 MPa for homogenization to obtain a homogenate. The pH value of the homogenate was adjusted to 5 with 1 M HCl and 1 M NaOH, 0.052 g (4 wt%) of cellulase (20,000 U / g) was added, and enzymolysis was carried out at 55°C for 2.5 h.

[0092] Cordyceps flower flavor peptide: After the dried cordyceps flower is crushed with a high-speed grinder, 40g is weighed, 600mL of pure water (solid-liquid ratio is 1:15) is added, and the colloid mill is used for pulping for 5min, and the high-pressure micro-jet homogenization pressure is set to 160MPa for homogenization to obtain a homogenate. The pH value of the homogenate is adjusted to 4.5 with 1M HCl and 1M NaOH, and 0.1g (4wt%) cellulase (20000U / g) is added, and enzymolysis is carried out at 50°C for 3h.

[0093] Boletus flavor peptide: After the dried boletus was crushed with a high-speed grinder, 40 g was weighed, 1000 mL of pure water (solid-liquid ratio was 1:25) was added, and the mixture was slurried with a colloid mill for 10 min, and the high-pressure microfluidization homogenization pressure was set to 180 MPa for homogenization to obtain a homogenate. Then, the pH value of the homogenate was adjusted to 4 with 1 M HCl and 1 M NaOH, 0.06 g (4 wt%) of cellulase (20,000 U / g) was added, and enzymolysis was carried out at 55°C for 2.5 h.

[0094] Pleurotus eryngii flavor peptide: After the dried Pleurotus eryngii was crushed with a high-speed grinder, 40 g was weighed, 800 mL of pure water (solid-liquid ratio was 1:20) was added, and the pulp was slurried with a colloid mill for 8 min, and the high-pressure micro-jet homogenization pressure was set to 180 MPa for homogenization to obtain a homogenate. Then, the pH value of the homogenate was adjusted to 5 with 1 M HCl and 1 M NaOH, 0.076 g (4 wt%) of cellulase (20000 U / g) was added, and enzymolysis was carried out at 55 ° C for 2 h.

[0095] White jade mushroom flavor peptide: After the dried white jade mushroom is crushed with a high-speed grinder, 40g is weighed, 800mL of pure water (solid-liquid ratio is 1:20) is added, and the pulp is slurried with a colloid mill for 10min, and the high-pressure micro-jet homogenization pressure is set to 180MPa for homogenization to obtain a homogenate. Then, the pH value of the homogenate is adjusted to 4 with 1M HCl and 1M NaOH, and 0.076g (4wt%) of cellulase (20000U / g) is added, and enzymolysis is carried out at 50°C for 2h.

[0096] Comparative Example 2

[0097] The flavor peptides of Lentinus edodes, Agrocybe oleraceus, Cordyceps sinensis, Boletus edulis, Pleurotus eryngii and Pleurotus eryngii were prepared by water extraction method. The specific steps were exactly the same as those of Examples 1 to 6, except that step (1) was water extraction method. The specific steps were as follows:

[0098] Shiitake mushroom flavor peptide: After the dried shiitake mushrooms are crushed with a high-speed grinder, 40 g are weighed, 800 mL of pure water (the solid-liquid ratio is 1:20) is added, and the mixture is boiled at 40°C for 3 h using a heat-collecting constant temperature heating magnetic stirrer to obtain a homogenate.

[0099] Agrocybe agrocybe flavor peptide: After the dried Agrocybe agrocybe was crushed with a high-speed grinder, 40 g was weighed, 1200 mL of pure water (solid-liquid ratio was 1:30) was added, and the mixture was boiled at 40°C for 3 h using a heat-collecting constant temperature heating magnetic stirrer to obtain a homogenate.

[0100] Cordyceps flower flavor peptide: After the dried cordyceps flower is crushed with a high-speed grinder, 40 g is weighed, 600 mL of pure water (solid-liquid ratio is 1:15) is added, and the mixture is boiled at 40°C for 2.5 h using a heat-collecting constant temperature heating magnetic stirrer to obtain a homogenate.

[0101] Boletus flavor peptide: After the dried boletus is crushed with a high-speed grinder, 40 g is weighed, 1000 mL of pure water (solid-liquid ratio is 1:25) is added, and the mixture is boiled at 40°C for 3 h using a heat-collecting constant temperature heating magnetic stirrer to obtain a homogenate.

[0102] Pleurotus eryngii flavor peptide: After the dried Pleurotus eryngii was crushed with a high-speed grinder, 40 g was weighed, 800 mL of pure water (solid-liquid ratio was 1:20) was added, and the mixture was boiled at 40°C for 3 h using a heat-collecting constant temperature heating magnetic stirrer to obtain a homogenate.

[0103] White jade mushroom flavor peptide: After the dry white jade mushroom is crushed with a high-speed grinder, 40 g is weighed, 800 mL of pure water (the solid-liquid ratio is 1:20) is added, and the mixture is boiled at 40°C for 2 h using a heat-collecting constant temperature heating magnetic stirrer to obtain a homogenate.

[0104] Comparative Example 3

[0105] The cellulose enzymatic hydrolysis method was used to prepare flavor peptides of shiitake mushroom, tea tree mushroom, cordyceps flower, boletus, king oyster mushroom and white jade mushroom. The specific methods were exactly the same as those in Examples 1 to 6, except that in step (1), the edible fungi were directly added with cellulase after being crushed by a high-speed crusher. The addition conditions, addition amount and enzymatic hydrolysis time of the cellulase in each edible fungus were exactly the same as those in Comparative Example 1.

[0106] Comparative Example 4

[0107] The ultrafine grinding method was used to prepare flavor peptides of shiitake mushroom, tea tree mushroom, cordyceps flower, boletus, king oyster mushroom and white jade mushroom. The specific methods were exactly the same as those in Examples 1 to 6, except that step (1) was changed to using an MFC-35B ultrafine grinder for grinding for 0.17h.

[0108] Experimental Test 1

[0109] The energy consumption ratios of flavor peptides of shiitake mushroom, Agrocybe edulis, Cordyceps sinensis, Boletus edulis, Pleurotus eryngii, and Pleurotus eryngii prepared by different preparation methods in Examples 1 to 6 and Comparative Examples 1 to 4 were investigated, and the results are shown in Table 1.

[0110] Table 1

[0111]

[0112]

[0113] As shown in Table 1, the energy consumption of Examples 1 to 6 (colloid mill-high pressure microfluidizer) is the lowest at 2433.6 ± 487kJ / h, and its soluble nitrogen release rate can be as high as 81.66 ± 0.96%, and the material loss rate reached 2.37±0.14%, while the energy consumption of comparative example 2 (colloid mill-high pressure microfluidizer + cellulose enzymatic hydrolysis) was the highest, reaching 18633.6±3726.13 kJ / h, and its soluble nitrogen release rate had no significant difference from that of the embodiment, and the working time was also extended by nearly 3 hours. Therefore, the combination of colloid mill-high pressure microfluidizer can omit the step of cellulose enzymatic hydrolysis and wall breaking.

[0114] From the working time point of view, Examples 1 to 6 (colloid mill-high pressure microfluidizer) and Comparative Example 4 (ultrafine grinding) are all better than Comparative Examples 1, 2, and 3. Although Comparative Example 4 (ultrafine grinding) has the shortest working time, its material loss rate is the highest at 15.48±0.26%; Comparative Example 2 (water extraction method) has lower energy consumption than Comparative Example 3 (cellulose enzymatic hydrolysis), but its soluble nitrogen release rate is the lowest at 46.57±1.21%, and there is no significant difference in the material loss rates of the two.

[0115] The particle sizes of the embodiments and comparative examples 1 to 4 prepared by different methods were investigated, and the results are shown in Table 2.

[0116] Table 2

[0117]

[0118]

[0119] As shown in Table 2, the particle size of edible fungi treated with colloid mill alone is in the range of 30-50 μm, ultrafine grinding can make the particle size of edible fungi reach 10-30 μm, and the combined treatment of colloid mill and high-pressure microjet homogenization can make the particle size of edible fungi reach 400-600 nm, thereby greatly improving the wall breaking efficiency.

[0120] Experimental Test Three

[0121] The sensory evaluation of the flavor peptides prepared in Examples 1 to 6 was performed as follows:

[0122] Sensory evaluation was performed using six aspects: color, aroma, bitterness, saltiness, umami, and taste richness. Ten (5 females and 5 males, aged 20-25 years) trained panelists from Sichuan Agricultural University were tested to evaluate the presented samples using a sensory standard rating scale (see Table 3), which was rated as 8-10 points-excellent, 6-8 points-very good, 4-6 points-good, 2-4 points-average, and 0-2 points-poor. The final score of different items (color, aroma, bitterness, saltiness, umami, and taste richness) was the average calculation of the ten scores. Before the sensory evaluation, sensory evaluation standards were prepared: the evaluation standards for sourness, sweetness, saltiness, bitterness, and umami were citric acid, sucrose, sodium chloride, L-isoleucine, and monosodium glutamate, with concentrations of 0.08%, 1%, 0.35%, 0.25%, and 0.35%, respectively. The standard at this concentration was rated 5 points. A score of 0-5 points represented a lower taste intensity than the standard, and 5-10 points represented a higher taste intensity than the standard. Sensory personnel were trained to be familiar with the use of the rating method, the terminology of each attribute, and the sensory characteristics of edible fungus flavor peptides. Each sample was repeated three times by the entire group. The results are as follows: Figure 1 shown.

[0123] Table 3

[0124]

[0125] Therefore, the present invention uses the colloid mill-high pressure microfluidizer combined technology to break and dissolve the edible fungi cells, which can make the material particle size reach the nanometer level, making the protein inside it easier to be degraded by enzymes, thereby significantly improving the enzymatic hydrolysis efficiency and omitting the traditional first step of cellulase enzymatic hydrolysis and cellulase wall breaking; the prepared flavor peptide has better quality, flavor and taste; no chemical reagents or solvents need to be added, reducing pollution to the environment. At the same time, the energy consumption required by the technology is relatively low, with an energy consumption of 2433.6 ± 487kJ / h, the soluble nitrogen release rate is as high as 81.66±0.96%, and the material loss rate is 2.37±0.14%, which helps to reduce production costs and meet the requirements of sustainable development.

[0126] Finally, it should be noted that the above embodiments are only used to illustrate the technical solution of the present invention rather than to limit it. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that they can still modify or replace the technical solution of the present invention with equivalents, and these modifications or equivalent replacements cannot cause the modified technical solution to deviate from the spirit and scope of the technical solution of the present invention.

Claims

1. A method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis, It is characterized in that Here are the steps: 1) After the dried edible fungi are crushed, water 10-30 times the mass of the dried powder is added, and the mixture is beaten with a colloid mill for 5-10 minutes, and then homogenized with a high-pressure microfluidizer to obtain an initial homogenate of the edible fungi for later use; 2) adjusting the pH value, adding flavor protease for enzymatic hydrolysis, and then killing the enzyme in boiling water; 3) adjusting the pH value again, adding xylose and lysine, and performing the Maillard reaction; 4) cooling the obtained edible fungus Maillard liquid to room temperature, and filtering out the edible fungus residue to obtain a supernatant; 5) pre-filtering the edible fungus Maillard liquid with an organic filter membrane, and then passing the edible fungus Maillard liquid through ultrafiltration membranes with molecular weight cutoffs of >10 kDa, 5k-10 kDa, 3k-5 kDa, and <3 kDa to obtain four components with different molecular weights; 6) spray drying the filtrate to obtain edible fungus flavor peptides of four components, namely, >10 kDa, 5k-10 kDa, 3k-5 kDa and <3 kDa, and performing sensory evaluation on the flavor peptides of different components; 7) Select the target peptide, and package the obtained target peptide using a heat shrink film packaging machine at room temperature.

2. The method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis according to claim 1, Features: Step 1) the pitch gap of the colloid mill is 100-200 μm, and the pressure parameter of the high-pressure microfluidizer is 160-200 MPa, so that the particle size of the edible fungi is not greater than 100 nm.

3. The method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis according to claim 1, Features: In step 2), the pH value is adjusted to 6-7, the amount of flavor protease added is 2-4wt% of the dry matter of the edible fungi, the enzymolysis time is 2-3h, and after the enzymolysis is completed, the enzyme is inactivated in boiling water for 15min.

4. The method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis according to claim 1, Features: In step 3), Maillard reaction is carried out, the reaction pH is 6.5-8, the reaction temperature is 90-120° C., the reaction time is 90-120 min, the amount of xylose added is 10-12 wt % of the dry matter of the edible fungus, and the amount of lysine added is 6-8 wt % of the dry matter of the edible fungus.

5. The method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis according to claim 1, Features: In step 4), four layers of gauze are used for filtering.

6. The method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis according to claim 1, Features: In step 5), the sample solution is pre-filtered using a 0.45 μm organic filter membrane.

7. The method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis according to claim 1, Features: In step 6), the spray drying parameters of the filtrate are as follows: air volume 70%, temperature 180° C., and feed rate 30%. The umami taste of flavor peptides of different components is evaluated using a sensory evaluation standard table.

8. Use of the method for preparing flavor peptides by colloid mill-high pressure microfluidization homogenization combined with assisted enzymatic hydrolysis as claimed in any one of claims 1 to 7 in the preparation of flavor peptides.

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